Understanding microbial adaptation within different natural environments remains a gap in knowledge in emerging infectious disease research and is particularly relevant toward improving public health. Tick-borne diseases continue to burden human health by continuously adapting in varying environments during the tick- mammalian infectious cycle. Relapsing fever (RF) spirochetes are one such pathogen, causing severe morbidity and mortality, yet little is known regarding the mechanisms involved with vector colonization and trans- mission. As RF spirochetes enter the tick they must adapt to disparate environments within the vector to continue their life-cycle, initially colonizing the midgut and thn the salivary glands, yet since RF spirochetes were first identified to cause human disease in 1905, two tick-associated proteins have been identified. To address this shortcoming, an in vivo system has been developed to study the complete tick- mammalian infectious cycle of Borrelia turicatae, a species of RF spirochete. A locus has been identified containing a series of genes that the spirochetes up-regulate during tick colonization. To date, this is the most comprehensive gene expression analysis of RF spirochetes within the arthropod vector. The genes are predicted to encode for outer membrane proteins (Omps), and given the importance of Omps in pathogen adaptation within the host and as vaccine targets, the proteins warrant further investigation. Thus, the identified genes are hypothesized to encode for Omps that are differentially produced within the midgut and salivary glands of the tick vector. The following aims will test the hypothesis: Aim 1: Determine the expression and surface localization of the predicted Omps in vitro. Rabbit immune serum generated against each putative recombinant Omp will be used to determine the surface localization and increased protein production when spirochetes are grown in tick-like conditions. Aim 2: Determine Omp production within the midgut and salivary glands of the tick. A method has been developed to cryopreserve and section ticks, clearly distinguishing spirochete populations within the midgut and salivary glands. Tick sections will be probed with rabbit serum generated against a given Omp to deter- mine protein production in the midgut and salivary glands. Completing this project will coincide with NIAID's mission to better understand and prevent disease. This project will result in the most comprehensive identification of surface proteins RF spirochete produce with- in the midgut and salivary glands, and guide mechanistic studies investigating protein function and the requirement of Omps during tick colonization and transmission. The results will also direct vaccine development against RF spirochetes, targeting bacterial Omps produced within the salivary glands, consequently neutralizing the bacteria as they enter the host.